Cholesterol metabolism altered and FGF21 levels high after pediatric liver transplantation despite normal serum lipids.

Liver transplantation (LT) predisposes to metabolic derangements and increases the risk for cardiovascular disease. We conducted a national cross-sectional study of all pediatric recipients who underwent LT between 1987 and 2007. We measured serum levels of noncholesterol sterols (surrogate markers of cholesterol synthesis and intestinal absorption) and fibroblast growth factor 21 (FGF21) in 49 patients (74% of survivors) at a median of 10 years posttransplant and in 93 controls matched for age and gender. Although serum cholesterol levels were similar in patients and controls, patients displayed increased whole-body synthesis and decreased intestinal absorption of cholesterol compared with controls (lathosterol to cholesterol ratio 129 ± 55 vs. 96 ± 41, respectively, p < 0.001; campesterol to cholesterol ratio 233 ± 91 vs. 316 ± 107, respectively; p < 0.001). Azathioprine (r =-0.383, p = 0.007) and low-dose methylpredisolone (r =-0.492, p < 0.001) were negatively associated with lathosterol/sitosterol ratio reflecting a favorable effect on cholesterol metabolism. FGF21 levels were higher in patients than in controls (248 pg/mL vs. 77 pg/mL, p < 0.001). In healthy controls, FGF21 was associated with cholesterol metabolism, an association missing in LT recipients. Normal serum lipids are achievable in long-term survivors of pediatric LT, but changes in cholesterol metabolism and increased FGF21 levels may explicate later cardiovascular risk.

Serum and lipoprotein sitostanol and non-cholesterol sterols after an acute dose of plant stanol ester on its long-term consumption.

PURPOSE: Chronic inhibition of cholesterol absorption with large doses of plant stanol esters (staest) alters profoundly cholesterol metabolism, but it is unknown how an acute inhibition with a large staest dose alters the postprandial serum and lipoprotein cholesterol precursor, plant sterol, and sitostanol contents. METHODS: Hypercholesterolemic subjects, randomly and double-blind divided into control (n = 18) and intervention groups (n = 20), consumed experimental diet without and with staest (plant stanols 8.8 g/day) for 10 weeks. Next morning after a fasting blood sample (0 h), the subjects had a breakfast without or with staest (4.5 g of plant stanols). Blood sampling was repeated 4 h later. Lipoproteins were separated with ultracentrifugation, and sterols were measured with gas-liquid chromatography. RESULTS: In 0-h chylomicrons and VLDL, plant sterols were lower in staest than in controls. Postprandially, cholestenol (cholesterol synthesis marker) was reduced in chylomicrons in staest compared with controls (-0.13 ± 0.04 µg/dL vs. 0.01 ± 0.08 µg/dL, P < 0.05). Staest decreased postprandially avenasterol in chylomicrons (P < 0.05 from 0 h). Sitostanol was high at 0 h by chronic staest in serum and VLDL but not in chylomicrons. Postprandial sitostanol was increased by staest in VLDL only. CONCLUSIONS: Chronic cholesterol absorption inhibition with large amount of plant stanol esters decreases plant sterols in triglyceride-rich lipoproteins. Acute plant stanol ester consumption increases sitostanol content in triglyceride-rich lipoproteins but suggests to decrease the risk of plant sterol and plant stanol accumulation into vascular wall by chylomicrons.

The role of serum non-cholesterol sterols as surrogate markers of absolute cholesterol synthesis and absorption.

AIMS: To study the whole-body cholesterol metabolism in man, cholesterol synthesis and absorption need to be measured. Because of the complicated methods of the measurements, new approaches were developed including the analysis of serum non-cholesterol sterols. In current lipidologic papers and even in intervention studies, serum non-cholesterol sterols are frequently used as surrogate markers of cholesterol metabolism without any validation to the absolute metabolic variables. The present review compares serum non-cholesterol sterols with absolute measurements of cholesterol synthesis and absorption in published papers to find out whether the serum markers are valid indicators of cholesterol metabolism in various conditions. DATA SYNTHESIS: During statin treatment, during interventions of dietary fat, and in type 2 diabetes the relative and absolute variables of cholesterol synthesis and absorption were frequently but not constantly correlated with each other. In some occasions, especially in subjects with apolipoprotein E3/4 and E4/4 phenotypes, the relative metabolic markers were even more sensitive than the absolute ones to reflect changes in cholesterol metabolism during dietary interventions. Even in general population at very high absorption the homeostasis of cholesterol metabolism is disturbed damaging the validity of the serum markers. CONCLUSIONS: It is worth using several instead of only one precursor and absorption sterol marker for making conclusions of altered synthesis or absorption of cholesterol, and even then the presence of at least some absolute measurement is valuable. During consumption of plant sterol-enriched diets and in situations of interfered cholesterol homeostasis the relative markers do not adequately reflect cholesterol metabolism. Accordingly, the validity of the relative markers of cholesterol metabolism should not be considered as self-evident.

Cholesterol, non-cholesterol sterols and bile acids in paediatric gallstones.

BACKGROUND: Depending on underlying aetiopathogenetic factors human gallstones contain various amounts of cholesterol, non-cholesterol sterols and bile acids, which have remained unexplored in paediatric gallstone patients. AIMS: To evaluate sterol and bile acids compositions of paediatric gallstones. PATIENTS AND METHODS: Study group included 21 consecutively cholecystectomised children. Gas-liquid chromatography was used to quantitate gallstone sterols and bile acids. Results were compared to adult gallstones (n=194). RESULTS: Cholesterol stones (n=9) had higher proportions of cholesterol and lathosterol, but lower those of lanosterol and phytosterols than pigment stones (n=12) (p<0.05 for each). Patients with gallstone cholesterol content over 70% were female. Gallstone cholesterol positively reflected body mass index and, in cholesterol stones-group, age (r=approximately +0.700, p<0.05). Three patients on parenteral nutrition had brown pigment stones consisting of high amounts of campesterol and sitosterol ranging 483-9303 microg/100 mg of stone. Pigment stones had 13-fold higher amount of bile acids than cholesterol stones (p<0.05). Black pigment stones contained approximately 3-fold higher phytosterol proportions, and pigment stones and cholesterol stones had approximately 43% lower proportions of deoxycholic acid than adults (p<0.05). CONCLUSION: Gallstones in patients on parenteral nutrition are rich in phytosterols. With respect to gallstone sterols, gallstone disease of adolescent girls resembles that of adults. Composition of bile acids in paediatric gallstones is different from adults.

Applicability of non-cholesterol sterols in predicting response in cholesterol metabolism to simvastatin and fluvastatin treatment among hypercholesterolemic men.

BACKGROUND AND AIMS: We hypothesized that (I) certain features in cholesterol metabolism at baseline could predict a response to statins, (II) good and poor responders to statins have a differential profile of serum and fecal sterols and (III) serum non-cholesterol sterols reflect cholesterol metabolism on statins. METHODS AND RESULTS: We examined serum lipids, serum and fecal cholesterol, cholesterol precursors, cholestanol and phytosterols and cholesterol metabolism among 20 hypercholesterolemic men at baseline and on 16-wk simvastatin/fluvastatin treatment. At baseline, the mean of serum cholestanol/cholesterol was 11% lower but those of lathosterol/cholesterol, lathosterol/cholestanol, desmosterol/cholesterol, desmosterol/cholestanol were 36-65% higher among good than poor responders (p<0.05 for each). On statins, reductions in ratios of serum precursor sterols and increases of absorption sterols were 1.8-2.9 times higher among good than poor responders (p<0.05 for each). In the whole study group, changes from baseline values of lathosterol/cholestanol were related to those of cholesterol and LDL-C in serum (r=+0.513 and +0.451, p=0.021 and 0.046, respectively). Serum lathosterol ratios to cholesterol, cholestanol and sitosterol consistently reflected a ratio of cholesterol synthesis (mg/d/kg)/fractional cholesterol absorption (%) (r-range +0.456 to +0.727, p<0.05 for each). CONCLUSIONS: Low serum baseline ratios to cholesterol of lathosterol, cholestenol and desmosterol, but a high ratio of cholestanol predicted a poor response to statins. Good responders were characterized by more profound reductions of serum and fecal (lathosterol) precursor sterols and increases of serum absorption marker sterol ratios on statins. Serum surrogate sterol markers of cholesterol metabolism were applicable in evaluating cholesterol absorption and synthesis also on statins.

Development of trunk asymmetry in a cohort of children ages 11 to 22 years.

STUDY DESIGN: A cohort study with a follow-up period of 11 years. OBJECTIVES: To study the growth of the spine with a focus on the development of trunk asymmetry and scoliosis. SUMMARY OF BACKGROUND DATA: Trunk asymmetry, a common phenomenon at adolescence, can be considered the clinical expression of scoliosis. The importance of the pubertal growth spurt has been stressed in the natural history of scoliosis. However, no cohort studies have focused on the ascending and descending phase of the spine's peak growth and the development of trunk asymmetry. METHODS: The cohort consisted of all the fourth-grade school children in the Western school district of Helsinki, Finland, in the spring of 1986. These 1060 children (515 girls and 545 boys), from the average age of 11 to 14 years, were invited to undergo annual examinations. The 855 children (80.7%) who had participated in the study at the age of 14 years were invited to a reexamination at the age of 22 years. This invitation was accepted by 430 (208 women and 222 men; 54%) of those invited. The forward bending test, the spinal pantography, and the anthropometric measurements were carried out by the same author (M.N.) throughout this study. RESULTS: At 22 years of age, 30% of the adults were found to be symmetric, with a hump less than 4 mm in the forward bending test, whereas 51% had a hump of 4 to 9 mm, and 19% had a hump 10 mm or larger (major asymmetry). The directional asymmetry of trunk surface, a skew to the right at the thoracic level and to the left at the lumbar level at puberty, remained constant at adult age. The prevalence of major trunk asymmetry at adult age was the same in both women and men, in contrast to the female predominance at puberty in this cohort. There were close correlations in the degrees of thoracic and lumbar asymmetry between puberty and adult ages. CONCLUSIONS: The shape of the back develops mainly during the pubertal growth spurt at ages 12 to 14 years in girls and boys. Trunk asymmetry (and mild scoliosis) seems as prevalent in young adult women as in men, although at puberty idiopathic scoliosis was twice as prevalent among girls as among boys in this cohort.

Smith-Lemli-Opitz syndrome and other sterol disorders among Finns with developmental disabilities.

Smith-Lemli-Opitz syndrome (SLOS) is an inherited disorder of cholesterol metabolism in which 7- and 8-dehydrocholesterols are accumulated in blood and tissues. Diagnosis of SLOS and other disorders in cholesterol metabolism (eg, cerebrotendinous xanthomatosis, phytosterolemia, desmosterolosis, and X-linked dominant Conradi-Hünermann-Happle syndrome) can be performed by gas-liquid chromatographic analysis of serum sterols. To elucidate their involvement in developmental disability, we evaluated serum sterols in two study groups: developmentally disabled subjects in long-term care (N = 322) and newborns and young children (N = 49) with features of SLOS in the Finnish population of 5 million. Only 1 SLOS case (type II) was found from among the 49 children. Seven additional adult cases (type I) with a wide range of clinical features and the serum sterol abnormalities characteristic of SLOS were detected from among the developmentally disabled subjects. The frequency of SLOS in the latter group was relatively high (7 in 322). No other hereditary sterol disorders were found, but two subgroups with low serum cholesterol precursor sterols and high serum plant sterols were identified. Several subjects, including the 7 SLOS patients, used ample medication and had abnormalities in serum sterol concentrations. Thus, among the subjects taking melperone, a high serum delta8-cholestenol level suggests an interference by the drug with cholesterol synthesis. Our results emphasize the importance of analyzing the serum sterols of developmentally disabled subjects to diagnose SLOS and of finding putative undiagnosed disorders in sterol metabolism associated with these clinical conditions.

Expression of histidine decarboxylase and cellular histamine-like immunoreactivity in rat embryogenesis.

In this study we investigated the developmental expression of histidine decarboxylase (HDC) mRNA and the distribution of histamine-immunoreactive (histamine-ir) cells in the rat embryonic tissues. We applied Northern blot analysis, in situ hybridization with synthetic oligonucleotide probes complementary to the rat HDC cDNA, and indirect histamine immunocytochemistry. Northern blot analysis revealed the appearance of a major (2.6 KB) HDC mRNA species in liver on embryonic Day 14. Its hybridization level peaked on Day E18, when two minor (1.6 and 3.5 KB) mRNA species were also present. During the periparturition period, a rapid decrease in HDC RNA was apparent, as the 2.6 KB mRNA species was expressed at a low level on postnatal Day P1. The embryonic liver expressed HDC on days E14-E20. On days E18 and E20, the periosteum and the epiphyseal growth plates of the endochondrally ossificating bones, and some striated muscle cells, showed hybridization signal for HDC. Histamine immunoreactivity was detected in many epithelial and neuronal cell types during embryogenesis. An intense histamine immunoreaction appeared first in essentially all cells of the liver parenchyma on day E12. This parenchymal histamine immunoreactivity disappeared by birth, after which this immunofluorescence in liver was restricted to a few scattered mast cells until adulthood. Some neurons in the peripheral sensory, sympathetic and cranial nerve ganglia were histamine-immunoreactive from day E16 to birth. In addition, many immunoreactive nerve fibers were detected in the gastrointestinal muscularis externa, mesentery, salivary glands, kidney, lung, and muscle tissue. We conclude that during rat embryogenesis histamine is produced and stored transiently by cells in liver, developing bone, and a few striated muscle cells, in addition to previously reported neurons in rat brain. Many peripheral neurons, epithelial cells, and mast cells display histamine immunoreactivity during rat embryogenesis but are devoid of detectable HDC mRNA with the current method. It remains possible that histamine is formed by another enzyme or is taken up from the extracellular space. The results support the concept that a significant proportion of histamine is formed and stored by embryonic cells other than mast cells.

Developmental patterns of histamine-like immunoreactivity in the mouse.

We studied the appearance and distribution of histamine (HA) during mouse embryogenesis, neonatal period, and adulthood using a specific rabbit HA antiserum and indirect immunofluorescence. HA first appeared on the Embryonic Day 13 (E13) in scattered mast cells in the gastrointestinal (GI) muscularis externa and liver. The splenic primordium contained a dense population of intensely HA-immunoreactive (HA-ir) cells from E13 on. From E15 to the birth, HA was detected in many embryonic cell types. On E15, the first HA-ir epithelial endocrine cells appeared in the oxyntic mucosa. In addition to the HA-ir cells in GI tract and liver, some nerve cells in ganglia of the peripheral nervous system (PNS), some fibers in spinal and cranial nerves, nerve fibers in mesenterium, and nerve plexuses of the gastrointestinal muscularis externa were HA-ir from E15 on. Occasional HA-ir nerve fibers were detected within the glandular epithelium of the oxyntic mucosa, pancreas, and salivary glands during late embryogenesis. During the same period, bright fluorescence was observed in cells of the kidney convoluted tubules and pancreatic islet cells. From E14 on, mast cells exhibiting bright fluorescence were scattered throughout the connective tissue of the fetus, and their number increased rapidly with age. Their density was especially high in subcutaneous connective tissue. Embryonic epidermal cells showed faint HA immunoreactivity. In musculoskeletal tissues, developing bone and occasional striated muscle cells exhibited HA immunoreactivity. Interestingly, most cells in liver showed transiently weak HA immunoreactivity during embryogenesis. In adult mouse, HA was stored only by scattered mast cells, oxyntic epithelial cells, and neurons in the tuberomamillary nucleus of the brain. The other HA-containing embryonic cells were negative for HA in adult mouse. In conclusion, HA immunoreactivity is widely distributed in epithelial, neuronal, and mast cells in various organs during mouse embryogenesis.

Histamine-storing cells in the oxyntic mucosa of the rat stomach: a transmission electron microscopic study employing fixation with carbodiimide.

We studied the distribution of histamine (HA) immunoreactivity in endocrine cells of the acid-producing mucosa in rat stomach with pre-embedding immunoelectron microscopy (IEM) using an antiserum against HA. Four fixation modifications were compared to optimize the ultrastructural morphology and staining pattern with the antisera produced against carbodiimide-conjugated HA. Fixation with 4% 1-ethyl-3(3-dimethyl-aminopropyl) carbodiimide (EDCDI) combined with both 4% paraformaldehyde and 0.1% glutaraldehyde gave superior results compared with EDCDI alone. Enterochromaffin-like (ECL) cells were easily distinguished from other endocrine cells in optimally fixed samples. The peroxidase end-product was distributed within the cytoplasm surrounding the vesicles of the ECL cells. ECL cells comprised about 75% of all endocrine cells, and about 90% of them were HA immunoreactive (HA-IR). No other HA-IR cell types were identified by EM in the basal half of the oxyntic region of rat gastric mucosa. The results suggest that a combination of EDCDI and aldehydes is suitable for IM demonstration of HA in cells. ECL cells from a predominant portion of endocrine cells in the oxyntic glands and may constitute the only significant non-mast cell store of HA in rat gastric mucosa.

Distribution of histamine in the rat kidney during pregnancy and development.

An antiserum against conjugated histamine and two oligonucleotide probes that detect the mRNA encoding L-histidine decarboxylase (HDC) involved in histamine synthesis were used to study the appearance of histamine and its location in the kidneys of fetal, newborn and young postnatal rats and in the kidneys of pregnant rats. On embryonic days 16 and 18 (E16 and E18), some HA-immunoreactive (HA-ir) cells were found within the largest S-shaped bodies. Histamine was found to appear rapidly between the 18th and 20th embryonic days in the convoluted tubules of the kidneys. On postnatal day 0 (P0), the distal convoluted tubules and collecting ducts exhibited bright fluorescence, the intensity of which decreased quickly so that it was faint on day P4 and absent at later stages. In kidneys of pregnant rats HA-ir was found in the epithelium of both the Bowman's capsule, collecting ducts and in a few cells within the tubules. Nonuniform HA-ir was also detected within glomeruli. No evidence for the presence of L-histidine decarboxylase mRNA in kidneys of fetuses or pregnant rats was seen. It is concluded that distinct structures in the developing rat kidney contain histamine during a period around birth from day E20 to day P4. In the pregnant rat, the epithelium that is in direct contact with the urine flow is immunoreactive for histamine from day 16 to 20 of pregnancy. The results suggest that histamine is not synthesized locally in the kidneys but rather originates from other tissues.

Ontogeny of histamine-immunoreactive cells in rat stomach.

An antiserum against hemocyanin-conjugated histamine was used to study the cellular stores of histamine in the stomach, especially the oxyntic mucosa, of fetal and early postnatal rats. Tissues were fixed in 4% 1-ethyl-3(3-dimethyl-aminopropyl) carbodiimide (EDC-DI) and standard immunofluorescence technique was used. Histamine was first detected on the 16th embryonic (E16) day when a few histamine-immunoreactive (HA-ir) cells and nerve fibers were observed in the muscular layer of the stomach wall. On day E18, HA-ir cells were visualized for the first time in the oxyntic mucosa of the stomach, and from that day on the number of such cells increased slowly initially and after day E20 more rapidly. At birth many of the HA-ir cells in the oxyntic mucosa possessed processes giving them a paracrine-like appearance typical of enterochromaffin-like cells (ECL cells). Only a very small number of the HA-ir cells represented metachromatically stained mast cells and were located in the submucosa. After birth, the number of HA-ir ECL cells increased steadily, until day 21 when the distribution and number was very similar to that of the adult. The results suggest that histamine-containing neurons and ECL cells appear in the stomach wall before birth, and that there are histamine-containing ECL cells in the mucosa and mast cells in the submucosa of the stomach wall at birth.

Distribution of histamine in developing rat tissues.

An antiserum against conjugated histamine was used to reveal the location and time of the appearance of the amine in different organs during the development of fetal and early postnatal rats. Tissues were fixed in 4% 1-ethyl-3(3-dimethyl-aminopropyl) carbodiimide and standard indirect immunofluorescence technique was used. Histamine-immunoreactivity (HA-ir) was first detected in the rat fetus on the 10th embryonic day (day E10) in the embryonic liver. The liver remained immunoreactive during the second half of the fetal life. In addition to some small HA-ir cells in the stomach wall and in the kidneys on day E16, the papillary muscles of the fetal heart also exhibited patchy immunofluorescence at this stage. On day E18, HA-ir cells appeared in many other organs including lungs, gastric mucosa and smooth muscle cells of the stomach wall, heart muscle and subcutaneous tissue. The distribution of HA-ir in fetal tissues was most extensive on day E20 when the kidney tubules and the skin also exhibited bright fluorescence. These results suggest that histamine is widely distributed during fetal development in different cell types.